The recurrent airway obstruction (RAO) in horses is etiologically characterized by an initially specific hyperresponsiveness to defined allergens and irritating stimuli, which becomes more and more unspecific when the disease progresses. Despite varying results obtained by different authors, it is generally assumed that an imbalance in the Th1/Th2 response with a shift to an excessive Th2 response and therefore a delayed, IgE-independent type IV immune response is the main immunologic process in horses with RAO.

The present study aimed to assess the specific immunomodulatory effect of inhalative administered cytosine-phosphate-guanine-oligodeoxynucleotides (CpG-ODN) over time. As a further goal, the study intended to verify the enhanced positive effect of a combined inhalation of A-class CpG-ODN and specific allergens. The inhalation of CpG-ODN should induce an immunomodulatory effect (Th1-Shift) and therefore alleviate RAO-symptoms quickly and in the longer term. Previous in vitro and animal-in vivo studies were able to proof this effect. In order to prevent a premature reduction of CpG-ODN by endogenous nucleases and furthermore to potentiate their effect in target cells, gelatin nanoparticles (GNP) were used as a transportation system for the cytosine-phosphate-guanine-oligodeoxynukleotides.

The study population contained 21 horses affected by RAO. Prior to the therapy, clinical and laboratory examinations of all horses were performed. The latter consisted of arterial blood gas analysis, a tracheobronchoscopy including a cytological analysis of the tracheobronchial secretion (TBS), an indirect measurement of the interpleural pressure and a functional in vitro test (FIT). Based on the results of these examinations, the horses were grouped into three levels of disease (mild, moderate, severe) and assigned by lot to two main groups. Ten horses received inhalation treatment with specific allergens and nanoparticle-bound CpG-ODN (group I), nine horses only received nanoparticle-bound CpG-ODN (group II) by inhalation. The inhalation protocol provided a total number of seven inhalations every second day. Additionally, within group I the amount of the specific allergens was gradually increased over the course of the inhalation treatment.

A second examination was performed after the last inhalation and a third examination six weeks after the end of the treatment protocol. Possible therapeutic success should be determined by clinical and endoscopic parameters, as well as tracheobronchial secretion cytology, both directly after the treatment and after a longer time span.

No local or systemic intolerance of the treatment occurred in any participants throughout the study. This suggests a very good tolerance of the inhalation protocol chosen for this study.

Neither the results of the first examination nor the results of the additional examinations led to statistically significant differences between the corresponding groups.

Both groups showed a marked improvement of the respiratory rate in rest directly after the last inhalation when compared to the first examination. This improvement was close to statistical significance. At the time of the third examination, group II showed a significant improvement, whereas the mean of group I reached almost physiological levels.

Concerning the auscultation results, group I showed a statistically significant decrease of the score-points in both follow-up examination. This statistical significance was reached by group II at the third examination. All values of the arterial blood gas analysis showed a distinct improvement in both groups after treatment. In the group treated with allergens (group I), these improvement of the parameters paO2 and A-aDO2 was significant at the third examination. In combination with the improved respiratory rate at rest, these results suggest that the inhalation treatment only with CpG-ODN as well as in combination with specific allergens is able to counteract the hypoxic metabolism caused by the abnormal gas exchange.

Due to the inhalation treatment both, a short- and long-term reduction of the interpleural pressure could be achieved for both groups. This suggests a reduction of the existing bronchoconstriction.

In group I both, the overall score and the endoscopy score significantly decreased directly after treatment as well as six weeks later. In some cases, the difference was even highly significant. Such a significant improvement was obtained for group II in the overall score and the tracheobronchial secret texture at the third examination.

Strikingly, the percentage of neutrophil granulocytes in TBS decreased distinctly in group I throughout the examinations, whereas in group II they remained almost constant.

Considering all findings, both group I and group II showed a marked improvement of all examination results. With the chosen treatment protocol in group I, a positive long-term effect on nearly all measured parameters was achieved. In group II 80% of the applied examination methods led to the same conclusion.

The presented method provides an alternative treatment option for horses with RAO to established treatment protocols. Regarding the examined parameters and the chosen treatment protocol, the combination of CpG-ODN and specific allergens seems to have no advantage over the use of CpG-ODN alone.

The results of this study should be reviewed in further studies involving larger patient groups as well as in dose-response studies. Especially the selection of the allergens, the amount of allergens and the administration interval must stand up to further investigations.